Capillary zone electrophoresis for monitoring r-DNA protein purification in multi-compartment electrolysers with immobiline membranes.
Isoforms of human monoclonal antibodies against the gp-41 of AIDS virus and of human recombinant superoxide dismutase have been purified to homogeneity by isoelectric focusing (IEF) in a multi-compartment electrolyser with isoelectric, immobiline membranes. This system allows the processing of large sample volumes and gram-scale protein loads and can resolve isoforms as close as 0.001 in pI difference. The purification progress was usually monitored by analytical IEF in immobilized pH gradients (IPG). Capillary zone electrophoresis (CZE) was applied to the monitoring of the content of each chamber of the electrolyser. CZE was found to be superior in terms of speed of analysis and quantification (but only by UV reading at 200-210 nm, i.e., in the region of the peptide bond) but, notwithstanding the millions of theoretical plates reported, was no match for the resolving power of IPGs, at least for protein analysis. When compared also with chromatofocusing, the resolving power decreases in the order IPG greater than CZE much greater than chromatofocusing.
Wenisch
E
E
Institute of Applied Microbiology, University of Agriculture and Foresty, Vienna, Austria.
Tauer
C
C
Jungbauer
A
A
Katinger
H
H
Faupel
M
M
Righetti
P G
PG
D000911:Antibodies, Monoclonal / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D004274:DNA, Recombinant / Q000235:genetics / Q000302:isolation & purification*; D004586:Electrophoresis / Q000379:methods*; D013006:Growth Hormone / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing; D007536:Isomerism; D008562:Membrane Glycoproteins / Q000276:immunology; D011506:Proteins / Q000235:genetics / Q000302:isolation & purification*; D015842:Serine Proteinase Inhibitors / Q000302:isolation & purification; D015843:Serpins; D013482:Superoxide Dismutase / Q000235:genetics / Q000302:isolation & purification
D000911:Antibodies, Monoclonal; D004274:DNA, Recombinant; D008562:Membrane Glycoproteins; D011506:Proteins; D015842:Serine Proteinase Inhibitors; D015843:Serpins; C053512:glycoprotein 41; D013006:Growth Hormone; D013482:Superoxide Dismutase